DNA Suspension Arrays: Silencing Discrete Artifacts for High-Sensitivity Applications

Detection of low frequency single nucleotide polymorphisms (SNPs) has important implications in early screening for tumorgenesis, genetic disorders and pathogen drug resistance. Nucleic acid arrays are a powerful tool for genome-scale SNP analysis, but detection of low-frequency SNPs in a mixed population on an array is problematic. We demonstrate a model assay for HIV-1 drug resistance mutations, wherein ligase discrimination products are collected on a suspension array. In developing this system, we discovered that signal from multiple polymorphisms was obscured by two discrete hybridization artifacts. Specifically: 1) tethering of unligated probes on the template DNA elicited false signal and 2) unpredictable probe secondary structures impaired probe capture and suppressed legitimate signal from the array. Two sets of oligonucleotides were used to disrupt these structures; one to displace unligated reporter labels from the bead-bound species and another to occupy sequences which interfered with array hybridization. This artifact silencing system resulted in a mean 21-fold increased sensitivity for 29 minority variants of 17 codons in our model assay for mutations most commonly associated with HIV-1 drug resistance. Furthermore, since the artifacts we characterized are not unique to our system, their specific inhibition might improve the quality of data from solid-state microarrays as well as from the growing number of multiple analyte suspension arrays relying on sequence-specific nucleic acid target capture

The trochlear isometric point is different in patients with recurrent patellar instability compared to controls: a radiographical study

Purpose: The purpose of the study was to investigate the theoretical isometric point based of the curve of the femoral groove and relating it to the origin of the MPFL femoral tunnel on lateral radiograph by comparing a patellar instability cohort with a control cohort. Methods: From a Patellar Instability database the radiographs of 40 consecutive patients were analysed to define Schöttle’s point, and the arc of the circle of the trochlear groove. A comparison population of 20 radiographs from comparable patients with tibiofemoral joint disorders was used as a control. The distance from Schöttle’s point to the most anterior part of the groove (extension) was also compared to the distance to the distal end of the roof of the notch (flexion). Results: The trochlea was circular in the controls but not the Patellofemoral Instability cohort where trochlear dysplasia is usually present. The difference between the extension and flexion length was a mean of − 2.0 ± 0.5 mm in the controls and + 6.0 ± 0.5 mm in the patellofemoral cohort. In neither cohort did the centre of the circle correspond to Schöttle’s point. The extension distance correlated with the boss height. Conclusions: The dysplastic trochlea is not circular and the centre of the best matched circle was different to the control trochleae which were circular. The circle centres did not correlate with Schöttle’s point for either cohort, and was more proximal in the Patellofemoral Instability cohort. Clinical relevance: For the MPFL to have equal tension throughout flexion within the groove, the length should not change. In normal knees the MPFL does not behave isometrically. The change in length, as measured from Schöttle’s point to the trochlea, was greater for patellofemoral instability patients explaining why an isolated MPFL reconstruction in the presence of severe trochlear dysplasia risks poor outcomes

Controls on gut phosphatisation : the trilobites from the Weeks Formation Lagerstätte (Cambrian; Utah)

Despite being internal organs, digestive structures are frequently preserved in Cambrian Lagerstätten. However, the reasons for their fossilisation and their biological implications remain to be thoroughly explored. This is particularly true with arthropods--typically the most diverse fossilised organisms in Cambrian ecosystems--where digestive structures represent an as-yet underexploited alternative to appendage morphology for inferences on their biology. Here we describe the phosphatised digestive structures of three trilobite species from the Cambrian Weeks Formation Lagerstätte (Utah). Their exquisite, three-dimensional preservation reveals unique details on trilobite internal anatomy, such as the position of the mouth and the absence of a differentiated crop. In addition, the presence of paired pygidial organs of an unknown function is reported for the first time. This exceptional material enables exploration of the relationships between gut phosphatisation and the biology of organisms. Indeed, soft-tissue preservation is unusual in these fossils as it is restricted to the digestive structures, which indicates that the gut played a central role in its own phosphatisation. We hypothesize that the gut provided a microenvironment where special conditions could develop and harboured a source of phosphorus. The fact that gut phosphatization has almost exclusively been observed in arthropods could be explained by their uncommon ability to store ions (including phosphorous) in their digestive tissues. However, in some specimens from the Weeks Formation, the phosphatisation extends to the entire digestive system, suggesting that trilobites might have had some biological particularities not observed in modern arthropods. We speculate that one of them might have been an increased capacity for ion storage in the gut tissues, related to the moulting of their heavily-mineralised carapace

Involvement of microbial mats in early fossilization by decay delay and formation of impressions and replicas of vertebrates and invertebrates

Microbial mats have been hypothesized to improve the persistence and the preservation of organic remains during fossilization processes. We test this hypothesis with long-term experiments (up to 5.5 years) using invertebrate and vertebrate corpses.Once placed on mats,the microbial community coats the corpses and forms a three-dimensional sarcophagus composed of microbial cells and exopolymeric substances (EPS). This coverage provides a template for i) moulding superficial features, resulting in negative impressions, and ii) generating replicas.The impressions of fly setulae, fish scales and frog skin verrucae are shaped mainly by small cells in an EPS matrix. Microbes also replicate delicate structures such as the three successive layers that compose a fish eye.The sarcophagus protects the body integrity, allowing the persistence of inner organs such as the ovaries and digestive apparatus in flies,the swim bladder and muscles in fish, and the bone marrow in frog legs.This study brings strong experimental evidence to the idea that mats favour metazoan fossilization by moulding, replicating and delaying decay. Rapid burial has classically been invoked as a mechanism to explain exceptional preservation. However, mats may play a similar role during early fossilization as they can preserve complex features for a long timeThis work, which is part of the research projects CGL2013-42643P and the research grant supporting M. Iniesto were funded by the Spanish Ministry of Economy and Competitiveness. The SEM facility at IMPMC was supported by Region Ile de France grant SESAME 2006 I-07-593/R, INSU-CNRS, INP-CNRS, and University Pierre et Marie Curie, Paris. SEM analyses performed for this study were supported by a grant from the Foundation Simone et Cino Del Duca (PI: K. Benzerara). Some SEM observations were also conducted at SIdI UAM (Madrid). Environmental SEM observations were performed at the MNCN (Madrid

Serum lipid responses to psyllium fiber: differences between pre- and post-menopausal, hypercholesterolemic women

<p>Abstract</p> <p>Background</p> <p>Cardiovascular disease is the leading cause of death in women and men. Psyllium, a soluble fiber has been known to reduce serum lipids. In this pilot study, we evaluated whether menopausal status would affect the serum lipid responses to psyllium fiber in women.</p> <p>Methods</p> <p>Eleven post-menopausal and eight pre-menopausal women with serum total cholesterol >200 mg/dL were included in the study. Subjects consumed their habitual diet and 15 g psyllium/d for 6 weeks. Psyllium was incorporated into cookies. Each cookie contained ≈5 g of psyllium fiber. Subjects ate one cookie in each meal.</p> <p>Results</p> <p>With psyllium fiber, total cholesterol concentration was significantly lower (≈5.2%, P < 0.05) in post-menopausal women but not in pre-menopausal women (≈1.3%). Also, there was a significant decrease in HDL-cholesterol in post-menopausal women (≈10.2%, P < 0.05). There were no significant changes observed in concentrations of LDL-cholesterol, triglycerides, apolipoprotein A1, and apolipoprotein B in both pre- and post-menopausal women with psyllium.</p> <p>Conclusion</p> <p>In this pilot study, post- and pre-menopausal, hypercholesterolemic women responded differently to psyllium fiber supplementation. Post-menopausal women would benefit from addition of psyllium to their diets in reducing the risk for heart diseases. The results of this study should be used with caution because the study was based on a small sample size.</p

Responses of marine benthic microalgae to elevated CO<inf>2</inf>

Increasing anthropogenic CO2 emissions to the atmosphere are causing a rise in pCO2 concentrations in the ocean surface and lowering pH. To predict the effects of these changes, we need to improve our understanding of the responses of marine primary producers since these drive biogeochemical cycles and profoundly affect the structure and function of benthic habitats. The effects of increasing CO2 levels on the colonisation of artificial substrata by microalgal assemblages (periphyton) were examined across a CO2 gradient off the volcanic island of Vulcano (NE Sicily). We show that periphyton communities altered significantly as CO2 concentrations increased. CO2 enrichment caused significant increases in chlorophyll a concentrations and in diatom abundance although we did not detect any changes in cyanobacteria. SEM analysis revealed major shifts in diatom assemblage composition as CO2 levels increased. The responses of benthic microalgae to rising anthropogenic CO2 emissions are likely to have significant ecological ramifications for coastal systems. © 2011 Springer-Verlag

Abnormal phospholipids distribution in the prefrontal cortex from a patient with schizophrenia revealed by matrix-assisted laser desorption/ionization imaging mass spectrometry

Schizophrenia is one of the major psychiatric disorders, and lipids have focused on the important roles in this disorder. In fact, lipids related to various functions in the brain. Previous studies have indicated that phospholipids, particularly ones containing polyunsaturated fatty acyl residues, are deficient in postmortem brains from patients with schizophrenia. However, due to the difficulties in handling human postmortem brains, particularly the large size and complex structures of the human brain, there is little agreement regarding the qualitative and quantitative abnormalities of phospholipids in brains from patients with schizophrenia, particularly if corresponding brain regions are not used. In this study, to overcome these problems, we employed matrix-assisted laser desorption/ionization imaging mass spectrometry (IMS), enabling direct microregion analysis of phospholipids in the postmortem brain of a patient with schizophrenia via brain sections prepared on glass slides. With integration of traditional histochemical examination, we could analyze regions of interest in the brain at the micrometric level. We found abnormal phospholipid distributions within internal brain structures, namely, the frontal cortex and occipital cortex. IMS revealed abnormal distributions of phosphatidylcholine molecular species particularly in the cortical layer of frontal cortex region. In addition, the combined use of liquid chromatography/electrospray ionization tandem mass spectrometry strengthened the capability for identification of numerous lipid molecular species. Our results are expected to further elucidate various metabolic processes in the neural system